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PubMed2026

Nucleic acid amplification testing and genome sequencing for WHO priority viruses in Africa: a scoping review.

Emerging viruses continue to pose serious public health threats across Africa, with recurrent outbreaks exposing gaps in diagnostics and surveillance systems. Nucleic acid amplification tests (NAATs) and genome sequencing are increasingly important for diagnostics and outbreak responses; however, their routine implementation is fragmented. This scoping review examines NAATs and genome sequencing technologies for viral detection and surveillance in Africa from 2019 to 2024, mapped to the 2024 updated WHO R&D Blueprint for Epidemics pathogen priority list. We identified 117 studies from 34 African countries reporting applications across 20 virus families, including ten designated as priorities by WHO. PCR-based assays were the most frequently reported NAATs. Illumina platforms predominated sequencing, and Oxford Nanopore Technologies were commonly used in outbreak investigations. Genome sequencing applied to priority viruses was largely reactive. NAAT-capable mobile laboratories were reported in 13 countries. Our findings underscore the need for proactive integration of NAATs into diagnostic and surveillance systems to strengthen decentralised testing, sustain genomic surveillance beyond outbreak periods, and improve early detection and preparedness for viral threats.

Journal Article↗

PubMed2026

Assessing the readiness of Oxford Nanopore sequencing for clinical genomics applications.

Long-read sequencing (LRS) technologies, namely, Oxford Nanopore Technologies (ONT) and Pacific Biosciences (PacBio), have emerged as promising solutions to overcome the limitations of short-read sequencing (SRS). Nevertheless, the still higher sequencing error rates compared with SRS, need for customized pipelines, rapidly updating software, and incipient scalability continue to present challenges for adopting ONT in standard clinical practice. Here we assess the performance of ONT (R9 and R10 chemistries) in comparison to Illumina and MGI across 17 well-characterized reference samples with 11 clinical variants representing nine different genetic diseases. To enable this, we have implemented a production-ready pipeline including SNV, indel, STR, SV, and CNV detection, alongside reporting key summary metrics to ensure high-quality data at the production sequencing level. Our results show high accuracy of ONT across SNVs (F-score 0.978-0.983) and SVs (F-score = 0.75) but still weaknesses across indels (F-score 0.659-0.758). However, we highlight that ONT accurately detected all four pathogenic indels as well as the performance improvement in exons and with the newer R10 chemistry. We further demonstrated the importance of long reads to detect clinically impactful variants such as a FMR1 pathogenic expansion, often misclassified by SRS as being in the premutation range. Our multiplatform analysis and Sanger validation uncovered a 1 bp error in the Coriell annotation for a cystic fibrosis-causing indel in GM07829. This work underscores the growing readiness of ONT for clinical applications, highlighting both its advancements and its potential for broader adoption in clinical genomics and large-scale operations.

Humans↗

PubMed2025

Molecular landscape of methicillin-resistant Staphylococcus aureus strains in clinical infections from hospitals in Lagos, Nigeria.

BACKGROUND AND OBJECTIVES: Multidrug-resistant Staphylococcus aureus (MRSA) accounts for a significant proportion of antimicrobial resistance (AMR)-associated infections worldwide. This study investigated the molecular profile of MRSA in Nigeria, providing valuable genomic data to fill existing knowledge gaps and highlighting its importance in the context of the global AMR crisis. METHODS: A total of 107 isolates were obtained from patient samples, including wound swabs/pus (65 isolates, 60.7%), blood cultures (16 isolates, 15%), urine/urinary catheter (8 isolates, 7.5%) and other sources. Species identification was performed using MALDI-TOF, and antimicrobial susceptibility testing was performed using the VITEK®2 system. Genomic DNA was extracted and subjected to whole-genome sequencing using short-read Illumina technology. In addition, a subset of isolates underwent long-read sequencing using Oxford Nanopore technology. RESULTS: Among the 107 isolates, 63 (59%) were identified as MRSA, with 58 (92%) carrying the mecA gene. The MRSA isolates exhibited high resistance to non-β-lactam antibiotics, particularly trimethoprim/sulfamethoxazole (95.3%), erythromycin (76.6%), gentamicin (71.4%) and quinolones (69.8%). The most prevalent MRSA belonged to the Bengal Bay clone [t657/ST772/Staphylococcal Cassette Chromosome mec (SCCmec) V(5C2)/Panton-Valentine leukocidin (PVL) + methicillin-susceptible Staphylococcus aureus (MRSA)], followed by t4690/ST152/SCCmec Vc(5C2&5)/PVL + MRSA and ST8 (t008, n = 1; t064, n = 4)/SCCmec Vc(5C2&5). Phylogenetic analysis suggests both community/associated transmission and possible importation of strains. CONCLUSIONS: This study highlights the significant burden of MRSA in Nigeria, with the high-risk Bengal Bay MRSA clone as the most common strain. The widespread resistance to non-β-lactam antibiotics underscores the urgent need for enhanced surveillance, infection control and antibiotic stewardship to mitigate its spread.

Journal Article↗

PubMed2026

Comparative evaluation of three high-molecular-weight DNA extraction kits for Oxford Nanopore sequencing of Clostridioides difficile and Clostridium perfringens.

UNLABELLED: Clostridioides difficile and Clostridium perfringens are Gram-positive, spore-forming anaerobic pathogens affecting humans and animals, for which genomic data have been mainly generated using short-read or hybrid sequencing approaches. In this study, we evaluated three commercial non-bead-beating DNA extraction kits designed for high-molecular-weight DNA recovery for Oxford Nanopore long-read whole-genome sequencing of two C. difficile and two C. perfringens strains, including one reference strain and one clinical or environmental isolate per species. Based on sequencing performance and kit ease of use, one kit was selected for additional sequencing of plasmid-carrying strains of both species. All three kits allowed correct identification of sequence types, toxin-encoding genes, and antimicrobial resistance determinants, confirming their suitability for clinical and epidemiological applications. However, the BT MasterPure Kit provided the highest DNA concentrations, longest fragment sizes, and superior read lengths and N50 values, particularly for C. difficile, achieving >100× coverage and enabling reliable circularization of chromosomes and plasmids, including a C. difficile metronidazole resistance plasmid and C. perfringens plasmids carrying toxin and antibiotic resistance genes. The other kits produced slightly lower DNA yields, resulting in shorter reads and reduced genome coverage for C. difficile, highlighting the challenge of extracting high-quality DNA from Gram-positive, spore-forming bacteria. Overall, this study provides practical guidance for selecting DNA extraction protocols optimized for Oxford Nanopore sequencing of C. difficile and C. perfringens, supporting high-quality genome assemblies and plasmid characterization and facilitating the routine genomic surveillance of clinically relevant spore-forming pathogens. IMPORTANCE: High-quality genomic data are essential for accurate characterization of Clostridioides difficile and Clostridium perfringens, two clinically and epidemiologically important Gram-positive, spore-forming pathogens. However, long-read sequencing performance can be strongly influenced by the choice of DNA extraction method, particularly for organisms with robust cell walls, where commonly used methods can lead to fragmented DNA. In this work, DNA of four strains was extracted using three commercial high-molecular-weight DNA extraction kits and sequenced using Oxford Nanopore Technologies. The best-performing kit was also evaluated using three additional strains known to harbor plasmids in order to assess its plasmid recovery efficiency. The results demonstrated successful plasmid recovery, circularization, and characterization. DNA extraction protocols optimized for Oxford Nanopore sequencing enable the rapid and cost-effective characterization of C. difficile and C. perfringens for genomic surveillance or outbreak investigations.

Clostridioides difficile↗

PubMed2025

Leveraging basecaller's move table to generate a lightweight k-mer model for nanopore sequencing analysis.

MOTIVATION: Nanopore sequencing by Oxford Nanopore Technologies (ONT) enables direct analysis of DNA and RNA by capturing raw electrical signals. Different nanopore chemistries have varied k-mer lengths, current levels, and standard deviations, which are stored in "k-mer models." In cases where official models are lacking or unsuitable for specific sequencing conditions, tailored k-mer models are crucial to ensure precise signal-to-sequence alignment, analysis and interpretation. The process of transforming raw signal data into nucleotide sequences, known as basecalling, is a fundamental step in nanopore sequencing. RESULTS: In this study, we leverage the move table produced by ONT's basecalling software to create a lightweight de novo k-mer model for RNA004 chemistry. We demonstrate the validity of our custom k-mer model by using it to guide signal-to-sequence alignment analysis, achieving high alignment rates (97.48%) compared to larger default models. Additionally, our 5-mer model exhibits similar performance as the default 9-mer models another analysis, such as detection of m6A RNA modifications. We provide our method, termed Poregen, as a generalizable approach for creation of custom, de novo k-mer models for nanopore signal data analysis. AVAILABILITY AND IMPLEMENTATION: Poregen is an open source package under an MIT license: https://github.com/hiruna72/poregen.

Nanopore Sequencing↗

PubMed2025

Degradation of ribosomal RNA during Plasmodium falciparum gametocytogenesis.

The life cycle of Plasmodium falciparum is characterized by complex regulatory changes that allow adaptation of the parasites to different environmental conditions, which are especially pronounced during transmission between the mammalian host and the insect vector. Previous studies have shown that P. falciparum uses three types of ribosomal RNAs (rRNA A, S1, and S2 types) at different stages of its life cycle. We used Oxford Nanopore Technologies direct RNA sequencing to investigate the dynamics of rRNA usage throughout the parasite's intraerythrocytic development, as well as in salivary gland sporozoites. Our study revealed a preponderance of A-type rRNAs during the intraerythrocytic cycle and gametocytogenesis, while S-type rRNAs slowly increase in abundance in mosquito stages starting 3 days post-infection. Salivary gland sporozoites showed an even proportion of all rRNA types. By examining the length distributions of rRNA molecules, we detected an extensive and specific degradation of rRNAs during gametocytogenesis, starting in stage II gametocytes and continuing until the final stages of gametocyte development. We hypothesize that rRNA degradation may be linked to the global translational repression and metabolic quiescence described in stage V gametocytes, similar to mechanisms observed in bacterial and eukaryotic stress responses.IMPORTANCEOur study uses Oxford Nanopore direct RNA sequencing of tightly synchronized blood-stage Plasmodium falciparum parasites to investigate the expression of ribosomal RNAs during asexual and sexual development. P. falciparum utilizes distinct types of rRNA during its development. However, due to the challenges of differentiating these highly similar molecules, their regulation and the mechanism underlying the switch between rRNA types remain unclear. We observe significant rRNA degradation in mature gametocytes, leading us to propose that this potentially leads to a reduced number of functional ribosomes when parasites become quiescent and translation is repressed.

Plasmodium falciparum↗

PubMed2026

First molecular detection and genomic characterization of West Nile virus associated with a naturally aborted sheep in Tunisia.

In September 2025, West Nile virus (WNV) RNA was detected for the first time in a sheep in Zaghouan Governorate, Tunisia. Following sheep abortions, fetal specimens were analyzed and one tested positive for WNV by RT-qPCR. The generated near-complete WNV genome sequence by Oxford Nanopore Technologies belonged to Lineage 1a, previously associated with locally reported human neuroinvasive cases in the same area. Amino acid analysis of the envelope (E) protein confirmed the presence of the conserved NYS glycosylation motif (E154-E156), a characteristic feature of Euro-Mediterranean Lineage 1 WNV strains. These findings further support the endemic circulation of WNV in Tunisia and underline its significance for both animal and public health.

Abortion↗

PubMed2026

Nanopore-based epigenomic profiling reveals the absence of widespread CpG methylation in the African swine fever virus genome.

DNA methylation is a critical epigenetic mechanism implicated in regulating replication and transcription in DNA viruses. However, the epigenetic landscape of African swine fever virus (ASFV), a large double-stranded DNA virus infecting pigs, remains controversial. Here, we systematically profiled the DNA methylome of the first ASFV strain isolated in Hong Kong (HK_NT_202103) using Oxford Nanopore Technologies (ONT) R10.4.1 sequencing. We employed a paired design: native whole-genome sequencing (WGS) against a methylation-free whole-genome amplification (WGA) control. Using conservative thresholds, we found no evidence of 5-methylcytosine (5mC), especially typical CpG methylation, across the viral genome. Importantly, clear CpG methylation signals were successfully detected in the host genome from WGS data, confirming the functionality of the workflow to detect 5mC at CG sites. While widespread 5mC seems absent, a small number of putative N6-methyladenine (6mA) loci were identified. A specific 6mA candidate exhibited raw ionic current disruptions and gene-level intersection with another ASFV isolate (CAS19-01/2019), although it lacked single-base consensus across different methylation callers or between the two isolates. Although our biological findings are restricted to a single isolate under specific experimental conditions, this study introduces a novel, highly rigorous ONT framework for viral epigenomics research. Furthermore, the absence of ASFV CpG methylation indicates that host CpG-depletion remains a viable strategy for viral metagenomic enrichment. Ultimately, our work offers a critical methodological baseline for ASFV surveillance and highlights the necessity of targeted experimental validation for rare viral modifications.

African Swine Fever Virus↗

PubMed2026

Radiation-resistant and desiccation-tolerant bacteria from the Chavara-Neendakara high background radiation area, india: phenotypic characterisation and genomic insights.

Radiation-resistant microorganisms that survive high doses of ionising radiation serve as valuable models for understanding stress adaptation; however, the genomic determinants underlying extreme radiation tolerance in bacteria from natural environments with high background radiation remain insufficiently characterised. Bacterial isolates from the Chavara-Neendakara HBRA (Kerala, India) were evaluated for desiccation tolerance, and the desiccation-resistant isolates were subsequently exposed to gamma irradiation (1-10 kGy) using a 60Co source. Isolates were identified through 16S rRNA sequencing, morphologically characterised by FE-SEM, and screened for antibiotic susceptibility. The highly radiation-resistant strain underwent whole-genome sequencing via Oxford Nanopore Technology, with De novo assembly, polishing, and genome annotation. Four bacterial isolates (Micrococcaceae and Paenibacillaceae) exhibited D10 values of 1-7 kGy, including one multidrug-resistant strain; no endospores were observed in the Paenibacillus isolate under the tested conditions. Paenibacillus sp. HBRA004 survived 10 kGy gamma radiation, exceeding all previously reported HBRA isolates by over fourfold. Its 5.0 Mbp genome (GC = 48.27%, ≥ 99% completeness) encodes five mechanistically independent DNA repair pathways; homologous recombination (recA, recN, radA), base excision repair (mutM, mutY, mutT), mismatch repair (mutL, mutS), nucleotide excision repair (uvrA, uvrB, uvrD), and non-homologous end joining (ku, ligD), alongside a redundant antioxidant network comprising triple-copy Fe/Mn-family superoxide dismutases and ahpC peroxiredoxin. A thioredoxin system (trxA, trxB, msrA) and manganese uptake via mntH may contribute to further layers of ROS defence. Their specific contribution to the HBRA004 phenotype remains to be experimentally and comparatively validated. These findings represent the first genomically characterised 10 kGy-resistant bacterial isolate from the Chavara-Neendakara HBRA, establishing a new benchmark for radiation tolerance within this ecologically significant environment. Pathway depth, gene copy amplification, and Mn/Fe homeostasis appear to be candidate mechanisms contributing to high-level radiation tolerance, consistent with patterns in other radiation-resistant taxa, though their contribution requires functional validation.

India↗

PubMed2026

Genomic signatures of host-range divergence in the generalist Beauveria bassiana and the specialist Beauveria brongniartii.

Entomopathogenic fungi of the genus Beauveria are widely used biological control agents that infect diverse insect hosts and can also associate with plants as rhizosphere colonizers and endophytes. Within this genus, Beauveria bassiana is a cosmopolitan generalist, whereas Beauveria brongniartii exhibits a narrower host range, primarily targeting soil-dwelling coleopteran larvae with limited evidence of plant colonization. To explore genomic differentiation associated with this ecological divergence, the commercially exploited B. brongniartii strain BIPESCO2 and B. bassiana ATHUM 4946 were sequenced using Oxford Nanopore technology, followed by comparative genomic analyses across multiple strains. Orthology identified species-specific gene families, although overall genome architecture and core gene content were highly conserved. The CAZyme repertoires were nearly identical, indicating retention of a versatile enzymatic toolkit supporting plant association, saprotrophy, and insect pathogenicity. In contrast, biosynthetic gene clusters displayed substantial variation, including structural remodeling of Beauveria-specific virulence-associated clusters and expansion of type I polyketide synthase clusters in B. brongniartii. Effector prediction revealed a conserved core of largely uncharacterized proteins alongside species-specific orthogroups enriched in adhesion-, immunity-, and cuticle-interaction domains. Together, these findings indicate that host-range divergence in Beauveria is associated with compartmentalized genomic differentiation, particularly in secondary metabolism and a limited subset of lineage-specific virulence factors, rather than in the conserved core infection machinery.

Beauveria↗

PubMed2026

Whole-genome sequence of the type strain and two field strains of Arsenicicoccus dermatophilus causing pododermatitis in greater flamingos (Phoenicopterus roseus).

The complete genome sequence of the type strain KM894/11T and two field strains of Arsenicicoccus dermatophilus (KM18/12; KM9/12) isolated from foot skin lesions of captive greater flamingos was determined using Oxford Nanopore and Illumina sequencing technologies. The genomes differ structurally by a large ~650 kb inverted chromosomal fragment and plasmid content.

dermatitis↗

PubMed2025

Detection and characterization of neonatal cytomegalovirus through nanopore sequencing using flongle flow cells: Pilot study in Philadelphia, Pennsylvania.

BACKGROUND: Cytomegalovirus (CMV) remains a significant infection in neonates and its early detection can aid with further treatment (antiviral, audiology). However, current diagnostics do not provide genetic information. OBJECTIVE: We explored the use of the portable and comprehensive sequencing method from Oxford Nanopore Technologies, utilizing low-cost Flongle flow cells to detect and perform sequence-level characterization of neonatal urine samples that tested positive for CMV by PCR. STUDY DESIGN: We performed a pilot study based on a retrospective cohort study of neonates who were positive for CMV by PCR, who were admitted at two birth hospitals in Philadelphia, PA. We leveraged deep and long-read sequencing results to analyze the reads in two forms: by comparing them against a reference-based strain and by reconstructing the genome through de novo assembly with phylogenetic tree analysis. RESULTS: We assayed seven clinical samples, including a positive and negative control sample, from newborns ranging from 23 weeks' gestation to term, with testing performed for microcephaly, hearing test results, small gestational age, and thrombocytopenia. Each sample showed multiple differences compared to the reference strain, and the phylogenetic tree analysis of the de novo assembly depicted the genetic diversity of the samples. CONCLUSION: This pilot study shows that nanopore sequencing with low-cost Flongle flow cells can detect and characterize CMV strains from clinical neonatal urine samples. This, coupled with current screening and diagnostic criteria, could further our genomic understanding of neonatal CMV, such as viral genome diversity, genotype-phenotype associations, and spread of strains.

Humans↗

PubMed2024

Sex differences in MAGEL2 gene promoter methylation in high functioning autism - trends from a pilot study using nanopore Cas9 targeted long read sequencing.

BACKGROUND: MAGEL2 is an autism susceptibility gene whose deficiency has been associated with autism-related behaviors in animal models and in syndromic human autism spectrum disorders (ASDs) such as Schaaf-Yang syndrome, but has not been studied in the broader autism spectrum. Given the capabilities of long-read sequencing technologies, this pilot study used a targeted nanopore sequencing approach to simultaneously examine MAGEL2 DNA sequence and methylation in adults with high-functioning autism (HFA) compared to neurotypical controls (NC). METHODS: Using DNA extracted from peripheral blood, Cas9-targeted nanopore DNA sequencing was used to analyze MAGEL2, including its entire regulatory construct (chr15:23639316-23651466), for sequence variation and 5-methyl-cytosine (5mC) modification in a cohort of adults with HFA compared to sex- and age-matched NC. Given the known sex differences in ASD and MAGEL2 KO animal models, results were further analyzed by sex. RESULTS: 20 adults with HFA (10 males, 10 females) and 20 NC were included. While there were no overall differences in MAGEL2 DNA sequence and 5mC modification between HFA and NC, we found a significant difference in MAGEL2 gene promoter methylation between males and females with HFA and NC of both sexes, with HFA males tending to show hypomethylation in a 300 bp long differentially methylated region (chr15:23647640-23647939) around the MAGEL2 transcription start site. CONCLUSIONS: In this pilot study utilizing nanopore Cas9 targeted DNA sequencing, significant sex-specific differences in MAGEL2 gene promoter methylation were identified in male adults with HFA in comparison to control groups, suggesting the potential for sex-specific epigenetic differences. However, further replication in larger cohorts is required to validate these findings.

Humans↗

PubMed2025

The chromosome-level genome of Stylosanthes guianensis provides insights into genome evolution and environmental adaptation.

Stylosanthes guianensis is a leguminous forage crop of significant economic importance, primarily distributed in tropical and subtropical regions. It exhibits strong adaptability to various stresses, yet the genetic basis underlying this trait remains unclear. In this study, we constructed the first chromosome-scale reference genome of S. guianensis using a combination of Nanopore and Hi-C sequencing technologies. The assembled genome size is 1254 Mb, with 10 pseudochromosomes. Using Nanopore full-length transcriptome data, we generated high-quality transcript-level gene annotations, identifying 36 585 gene models and 110 601 transcripts. The repetitive sequences in S. guianensis account for 79.16% of the genome, with the extensive expansion of Gypsy elements in long terminal repeats contributing to its genome size enlargement. Comparative genomic and transcriptomic analyses revealed that flavonoid metabolism plays a pivotal role in stress adaptation, providing new insights into the genetic basis of stress tolerance. Additionally, we generated whole-genome methylation profiles under cold treatment and control conditions, offering valuable data for future epigenomic research. These findings provide essential molecular resources for understanding stress resilience in S. guianensis and advancing its molecular breeding.

Genome, Plant↗

PubMed2026

Dissecting the relationship between haplotypes around ATXN2 CAG repeats and the number of CAA interruptions by long-read sequencing.

BACKGROUND: CAG repeat expansions in ATXN2 are implicated as risk factors for several neurological diseases, including spinocerebellar ataxia type 2 (SCA2) when >=33 CAG repeats are present, and amyotrophic lateral sclerosis (ALS) when 27-33 CAG repeats are present. However, how haplotypes around the repeats and CAA interruptions within the repeats are associated with disease phenotypes remains poorly understood. Previous studies on haplotypes around ATXN2 were limited to SNPs very close to the repeats (<5kb) or were based on statistical inference only. METHODS: Here, we used long-read sequencing on the Oxford Nanopore Technologies (ONT) platform to simultaneously infer haplotypes around ATXN2, the number of CAG repeats, and the number of CAA interruptions, along with NYGC ALS Consortium NGS dataset. We further sequenced 41 individuals (EUR = 39) with neurological diseases with intermediate repeats by ONT. RESULTS: We found that haplotypes around ATXN2 and the number of interruptions show ethnicity-specific and ALS-specific distribution. Three CAA interruptions are present at low prevalence (~1%) in control populations in multiple ancestry groups, but high prevalence (~55%) in ALS individuals with intermediate repeats. Furthermore, we examined 159 individuals with ALS (~90% European ancestry) with intermediate ATXN2 repeats and found a unique haplotype in ALS individuals with three CAA interruptions, which can be tagged by an SNV, rs148019457. We also validated that the rs148019457-G allele is only present in haplotypes with three CAA interruptions. CONCLUSIONS: In summary, our study shows that 3 CAA interruptions are rarely seen in healthy controls but are common in those with expanded ATXN2 CAG repeats who have neurological disorders, and that rs148019457 tags a specific haplotype with 3 CAA interruptions within expanded ATXN2 CAG repeats in individuals of European ancestry. These results have implications for the development of precision genomic medicine for neurological disorders, and the tag SNP may help identify those with interruptions from existing population genotyping data.

ATXN2↗

PubMed2025

Allele Level Sequencing of Killer Cell Immunoglobulin-Like Receptor Genes Using Oxford Nanopore Long Read Sequencing.

The human Killer cell Immunoglobulin-like Receptor (KIR) genes, found on chromosome 19, encode for cell surface protein receptors that, through interaction with their ligand, modulate the action of Natural Killer (NK) cells and some subsets of T lymphocytes. KIR genes exhibit extensive variation through variable gene content, copy number, and allele polymorphism. The combination of KIR genes and their ligands is implicated in various clinical settings including haematopoietic stem cell and solid organ transplant, and infectious disease progression. KIR gene content has been used in the selection of optimal stem cell donors with haplotype variations in recipient and donor giving differential clinical outcomes. With the introduction of massively parallel clonal next generation sequencing and single molecule long read third generation sequencing, allele level determination of KIR genotypes has become feasible. We describe a method for amplicon-based long read sequencing on the Oxford Nanopore Technologies platform that provides largely unambiguous allele level typing of KIR genes. The method was validated using DNA extracted from 48 10th International Histocompatibility Workshop (IHWS) cell lines with previously published allele level KIR genotypes and 176 Western Australian samples previously tested for the presence or absence of KIR genes. Our long-read sequencing method was able to accurately determine KIR alleles with an overall concordance of 97%-99% with the published data. Importantly, phasing ambiguity caused by the inability to phase heterozygous base positions over long stretches of gene sequence was resolved in several samples. Thus, our long read PCR sequencing strategy can be used to determine KIR genotypes at allele resolution level.

Humans↗

PubMed2026

Whole-genome sequences of the dwarf honey bee subgenus Micrapis: Apis andreniformis and Apis florea.

The Micrapis subgenus, which includes the black dwarf honey bee (Apis andreniformis) and the red dwarf honey bee (Apis florea), remains underrepresented in genomic studies despite its ecological significance. Here, we present high-quality de novo genome assemblies for both species, generated using a hybrid sequencing approach combining Oxford Nanopore Technologies long reads with Illumina short reads. The final assemblies are highly contiguous, with contig N50 values of 5.0 Mb (A. andreniformis) and 4.3 Mb (A. florea), representing a major improvement over the previously published A. florea genome. Genome completeness assessments indicate high quality, with BUSCO scores exceeding 98.5% using the Hymenoptera database and k-mer analyses supporting base-level accuracy. Repeat annotation revealed a relatively low repetitive sequence content (∼6%), consistent with other Apis species. Using RNA sequencing data, we annotated 12,189 genes for A. andreniformis and 12,207 genes for A. florea, with ∼98% completeness in predicted proteomes. These genome assemblies provide a valuable resource for comparative and functional genomic studies, with the potential to offer new insights into the genetic basis of dwarf honey bee adaptations.

Male↗

PubMed2025

Determining genotype and antimicrobial resistance of Salmonella Typhi in environmental samples by amplicon sequencing.

BACKGROUND: Estimates of the burden of typhoid fever due to Salmonella enterica serovar Typhi (S. Typhi) rely on data from clinical surveillance, which is rarely done in low income settings and is also limited by the poor sensitivity of the assays used and the reliance on health seeking by patients. Environmental surveillance for S. Typhi shed by symptomatic and asymptomatic individuals in wastewater offers a sensitive surveillance tool that could help to inform burden estimates. Sequencing S. Typhi direct from wastewater concentrates has the potential to identify circulating genotypes and associated antimicrobial resistance (AMR) genes, supporting public health interventions such as vaccination and antimicrobial usage. METHODOLOGY AND PRINCIPAL FINDINGS: We designed a multiplex targeted amplicon sequencing protocol for genotyping and determining AMR in S. Typhi from wastewater samples, targeting SNPs that identify genotypes of interest and both chromosomal and plasmid-borne AMR. PCR products were sequenced using the Oxford Nanopore Technologies (ONT) MinION, and genotypes and AMR identified using the GenoTyphi program. We tested this approach on samples from south India from both hospital outflow and wastewater collected from the community. All samples tested were suspected to be positive for S. Typhi following quantitative PCR for ttr, tviB, and staG gene targets. Out of 110 samples tested we were able to determine a genotype and/or AMR for 8. All samples that gave a genotype call suggested a genotype consistent with those found in clinical cases in India during the same time period and produced consensus sequences that clustered with S. Typhi when included in a phylogenetic tree. CONCLUSIONS: In this study, we provide proof of concept data for amplicon sequencing of S. Typhi in wastewater which with further optimisation could be used to complement clinical surveillance data or provide data on S. Typhi presence in the absence of clinical surveillance. This information can inform public health interventions, and the concept could be applied to other pathogens of interest for genotyping from environmental surveillance samples.

Salmonella typhi↗